Parkinson&#39;s disease treatment

ABSTRACT

Methods for treating a movement disorder by intracranial administration to a human patient of a therapeutically effective amount of a neurotoxin, such as a botulinum toxin type A.

CROSS REFERENCE

This application is a divisional of Ser. No. 09/596,306, filed Jun. 14,2000, now U.S. Pat. No. 6,306,403.

BACKGROUND

The present invention relates to methods for treating movementdisorders. In particular, the present invention relates to methods fortreating movement disorders by intracranial administration of aneurotoxin.

Movement Disorders

A movement disorder is a neurological disturbance that involves one ormore muscles or muscle groups. Movement disorders include Parkinson'sdisease, Huntington's Chorea, progressive supranuclear palsy, Wilson'sdisease, Tourette's syndrome, epilepsy, and various chronic tremors,tics and dystonias. Different clinically observed movement disorders canbe traced to the same or similar areas of the brain. For example,abnormalities of basal ganglia (a large cluster of cells deep in thehemispheres of the brain) are postulated as a causative factor indiverse movement disorders.

Tremors are characterized by abnormal, involuntary movements. Anessential tremor is maximal when the body part afflicted (often an armor hand) is being used, for example when attempts at writing or finecoordinated hand movements are made. Typical chemotherapy is use of thedrug propranolol (Inderal) which has the side effects of low bloodpressure and heart rate changes. A resting tremor is common inParkinson's disease and in syndromes with Parkinsonian features. Aresting tremor is maximal when the extremities are at rest. Often, whena patient attempts fine movement, such as reaching for a cup, the tremorsubsides. Systemic anticholinergic medications have been used with somesuccess.

Dystonias are involuntary movement disorders characterized by continuedmuscular contractions which can result in twisted contorted posturesinvolving the body or limbs. Causes of dystonia include biochemicalabnormalities, degenerative disorders, psychiatric dysfunction, toxins,drugs and central trauma. Thalamotomy and/or subthalamotomy or campotomyare currently the preferred neurosurgical procedures to treat dystonia,and are carried out with techniques and brain targets similar to thesurgical treatment of Parkinson's disease. Tasker R., Surgical Treatmentof the Dystonias, chapter 105, pages 1015-1032, in Gildenberg P. L. etal., Textbook of Stereotactic and Functional Neurosurgery, McGraw-Hill(1998).

Particular dystonias can include spasmodic torticollis, blepharospasmand writer's cramp. Spasmodic torticollis is a syndrome that usuallyaffects adults, and involves the involuntary turning of the neck to oneside. Some individuals may not even notice initially that the head andneck are turned. Blepharospasm is an involuntary movement which involvesintermittent forceful closure of the eyelids. Writers cramp is acramping abnormal posture which develops when one is writing, orperforming other actions with the hands. Symptoms may progress toinvolve the arm and shoulder.

Tic disorders (including Tourette's) are usually very rapid, short livedstereotyped repeated movements. The more common tics involve the motorsystems, or are vocal in nature. Motor tics often involve the eyelids,eyebrows or other facial muscles, as well as the upper limbs. Vocal ticsmay involve grunting, throat clearing, coughing or cursing. Individualswith tic disorders will often describe a strong urge to perform theparticular tic, and may actually feel a strong sense of pressurebuilding up inside of them if the action is not performed. For example,a motor tic that may involve the abrupt movement of one of the arms maybe controllable for a short period of time if the affected person sitson his hands; however, the almost irresistible urge to do the actionoften takes over and result in the tic action.

Tourette's syndrome is a tic disorder which often begins in childhood oradolescence and is much more common in males. There are both multiplemotor tics, as well as vocal tics present. The tics often change frominvolvement of one body part to another, and the disease often getsbetter and worse intermittently, with periods of almost minimalactivity, and other times when some patients have difficultyfunctioning. Other neurobehavioral difficulties often accompany thesyndrome. These include attention deficit hyperactivity disorder (ADHD)and obsessive-compulsive disorder. Treatment of most tic disordersemploys the use of medications that decrease the amount of dopamine inthe brain, such as dopamine antagonists. Unfortunately these drugs areassociated with side effects such as other movement disorders, includingParkinsonism (stiffness, slow movement and tremors). In addition toTourette's syndrome, tics may be associated with head injury, carbonmonoxide poisoning, stroke, drug use and mental retardation.

Progressive supranuclear palsy is a movement disorder in which patientshave significant difficulty moving their eyes vertically (up and down)initially, followed by all eye movements become limited(ophthalmoplegia). Patients can also develop dementia, rigidity,bradykinesia (slow movements) and a propensity for falls.

Huntington's chorea is a genetically inherited disorder that has bothneurological and psychiatric features. Most cases develop when peopleare in their forties or fifties, but early and late onset is alsopossible. The disease may begin with either the neurological or mentalstatus changes. The neurological symptoms may vary, but include chorea.Chorea (derived from a Greek word meaning, “to dance”) is a series ofmovements that is dance-like, jerky, brief, and moves from one part ofthe body to another. Clumsiness, fidgetiness and jumpiness may alsooccur. Facial movements, especially around the jaw, may occur. There isoften difficulty with walking and posture. The psychiatric symptoms maypresent as paranoia, confusion, or personality changes. As the diseaseprogresses, a significant dementia develops. MRI brain imaging may showatrophy (shrinkage) of a portion of the basal ganglia (involved inmovement) that is known as the caudate nucleus.

Wilson's disease is a disorder that involves the nervous system andliver function. The neurological problems include tremors,incoordination, falling, slurred speech, stiffness and seizures.Psychiatric problems can occur and patients can develop severe liverdamage if this affliction is untreated. Elevated copper andceruloplasmin levels are diagnostic.

Unfortunately, a movement disorder, including those set forth above, canbecome resistant to drug therapy. Drug resistant tremors can includeresting tremors, such as can occur in Parkinson's disease, and actiontremors, such as essential tremor, multiple sclerosis tremors, posttraumatic tremors, post hemiplegic tremors (post stroke spasticity),tremors associated with neuropathy, writing tremors and epilepsy.

Parkinson's Disease

Parkinson's disease is a movement disorder of increasing occurrence inaging populations. Parkinson's disease is a common disabling disease ofold age affecting about one percent of the population over the age of 60in the United States. The incidence of Parkinson's disease increaseswith age and the cumulative lifetime risk of an individual developingthe disease is about 1 in 40. Symptoms include pronounced tremor of theextremities, bradykinesia, rigidity and postural change. A perceivedpathophysiological cause of Parkinson's disease is progressivedestruction of dopamine producing cells in the basal ganglia whichcomprise the pars compartum of the substantia nigra, a basal nucleilocated in the brain stem. Loss of dopamineric neurons results in arelative excess of acetylcholine. Jellinger, K. A., Post Mortem Studiesin Parkinson's Disease—Is It Possible to Detect Brain Areas For SpecificSymptoms?, J Neural Transm 56 (Supp);1-29:1999.

Parkinson's disease is a progressive disorder which can begin with mildlimb stiffness and infrequent tremors and progress over a period of tenor more years to frequent tremors and memory impairment, touncontrollable tremors and dementia.

Drugs used to treat Parkinson's disease include L-dopa, selegiline,apomorphine and anticholinergics. L-dopa (levo-dihydroxy-phenylalanine)(sinemet) is a dopamine precursor which can cross the blood-brainbarrier and be converted to dopamine in the brain. Unfortunately, L-dopahas a short half life in the body and it is typical after long use (i.e.after about 4-5 years) for the effect of L-dopa to become sporadic andunpredictable, resulting in fluctuations in motor function, dyskinesiasand psychiatric side effects. Additionally, L-dopa can cause B vitamindeficiencies to arise.

Selegiline (Deprenyl, Eldepryl) has been used as an alternative toL-dopa, and acts by reducing the breakdown of dopamine in the brain.Unfortunately, Selegiline becomes ineffective after about nine months ofuse. Apomorphine, a dopamine receptor agonist, has been used to treatParkinson's disease, although is causes severe vomiting when used on itsown, as well as skin reactions, infection, drowsiness and somepsychiatric side effects.

Systemically administered anticholinergic drugs (such as benzhexol andorphenedrine) have also been used to treat Parkinson's disease and actby reducing the amount of acetylcholine produced in the brain andthereby redress the dopamine/acetylcholine imbalance present inParkinson's disease. Unfortunately, about 70% of patients takingsystemically administered anticholinergics develop seriousneuropsychiatric side effects, including hallucinations, as well asdyskinetic movements, and other effects resulting from wideanticholinergic distribution, including vision effects, difficultyswallowing, dry mouth and urine retention. See e.g. Playfer, J. R.,Parkinson's Disease, Postgrad Med J, 73;257-264:1997 and Nadeau, S. E.,Parkinson's Disease, J Am Ger Soc, 45;233-240:1997.

Before the introduction of L-dopa in 1969, stereotactic surgery offeredone of the few effective treatments for Parkinson's disease. Thesignificant known deficiencies and drawbacks associated with therapeuticdrugs to treat Parkinson's disease, including the long term limitationsof L-dopa therapy have led to renewed interest in neurosurgicalintervention. Unilateral stereotactic thalamotomy has proven to beeffective for controlling contralateral tremor and rigidity, but carriesa risk of hemiparesis. Bilateral thalamotomy carries an increased riskof speech and swallowing disorders resulting. Stereotactic pallidotomy,surgical ablation of part of the globus pallidus (a basal ganglia), hasalso be used with some success. Aside from surgical resection, highfrequency stimulating electrodes placed in the ventral intermedialisnucleus has been found to suppress abnormal movements in some cases. Avariety of techniques exist to permit precise location of a probe,including computed tomography and magnetic resonance imaging.Unfortunately, the akinesia, speech and gait disorder symptoms ofParkinson's disease are little helped by these surgical procedures, allof which result in destructive brain lesions.

Epilepsy

A seizure is a paroxysmal event due to abnormal, excessive,hypersynchronous discharges from an aggregate of central nervous systemneurons. Epilepsy describes a condition in which a person has recurrentseizures due to a chronic, underlying process. Among the many causes ofepilepsy there are various epilepsy syndromes in which the clinical andpathologic characteristics are distinctive and suggest a specificunderlying etiology. The prevalence of epilepsy has been estimated at 5to 10 people per 1000 population. Severe, penetrating head trauma isassociated with up to a 50% risk of leading to epilepsy. Other causes ofepilepsy include stroke, infection and genetic susceptibility.

Antiepileptic drug therapy is the mainstay of treatment for mostpatients with epilepsy and a variety of drugs have been used. See e.g.,Fauci, A. S. et al., Harrison's Principles of Internal Medicine,McGraw-Hill, 14^(th) Edition (1998), page 2321. Twenty percent ofpatients with epilepsy are resistant to drug therapy despite efforts tofind an effective combination of antiepileptic drugs. Surgery can thenbe an option. Video-EEC monitoring can be used to define the anatomiclocation of the seizure focus and to correlate the abnormalelectrophysiologic activity with behavioral manifestations of theseizure. Routine scalp or scalp-sphenoidal recordings are usuallysufficient for localization. A high resolution MRI scan is routinelyused to identify structural lesions. Functional Imaging studies such asSPECT and PET are adjunctive tests that can help verify the localizationof an apparent epileptogenic region with an anatomic abnormality. Oncethe presumed location of the seizure onset is identified, additionalstudies, including neuropsychological testing and the intracarotidamobarbital test (Wada's test) can be used to assess language and memorylocalization and to determine the possible functional consequences ofsurgical removal of the epileptogenic region. In some cases, the exactextent of the resection to be undertaken can be determined by performingcortical mapping at the time of the surgical procedure. This involveselectrophysiologic recordings and cortical stimulation of the awakepatient to identify the extent of epileptiform disturbances and thefunction of the cortical regions in questions.

The most common surgical procedure for patients with temporal lobeepilepsy involves resection of the anteromedial temporal lobe (temporallobectomy) or a more limited removal of the underlying hippocampus andamygdala. Focal seizures arising from extratemporal regions may besuppressed by a focal neocortical resection. Unfortunately, about 5% ofpatients can still develop clinically significant complications fromsurgery and about 30% of patients treated with temporal lobectomy willstill have seizures.

Focal epilepsy can involve almost any part of the brain and usuallyresults from a localized lesion of functional abnormality. One type offocal epilepsy is the psychomotor seizure. Current therapy includes useof an EEG to localize abnormal spiking waves originating in areas oforganic brain disease that predispose to focal epileptic attacks,followed by surgical excision of the focus to prevent future attacks.

Brain Motor Systems

Several areas of the cerebrum influence motor activity. Thus, lesion tothe motor cortex of the cerebrum, as can result from stoke, can removeinhibition of vestibular and reticular brain stem nuclei, which thenbecome spontaneously active and cause spasm of muscles influenced by,the now disinhibited, lower brain areas.

An accessory motor system of the cerebrum is the basal ganglia. Thebasal ganglia receives most input from and sends most of its signalsback to the cortex. The basal ganglia include the caudate nucleus,putamen, globus pallidus, substantia nigra (which includes the parscompacta) and subthalamic nucleus. Because abnormal signals from thebasal ganglia to the motor cortex cause most of the abnormalities inParkinson's disease, attempts have been made to treat Parkinson'sdisease by blocking these signals. Thus lesions have been made in theventrolateral and ventroanterior nuclei of the thalamus to block thefeedback circuit from the basal ganglia to the cortex. Additionally,pallidotomy, the surgical ablation of part of the globus pallidus, hasbeen used to effectively treat the motor disorders of Parkinson'sdisease.

Surgical intervention is believed to assist by interrupting a motoricpathway which, due to a dopaminergic deficiency, had pathologicallyinhibited the thalamus. The inhibited thalamus in turn understimulatescortical neuronal networks responsible for generating movement. Hence,surgery removes the thalamic inhibition and has been used in thetreatment of pharmacoresistant movement disorders. Speelman, J. D., etal., Thalamic Surgery and Tremor, Mov Dis 13(3);103-106:1998.

Intracranial lesions for the treatment of tremor and other parkinsoniansymptoms have been made to the globus pallidus and the ansalenticularis. Long term results of pallidotomy have sometimes beendisappointing. Positive results for the surgical arrest of tremor havebeen obtained by lesioning the following thalamic nuclei: (1) theventrointermedius (Vim) or ventral lateral posterior (VLp) nucleus; (2)ventrooralis anterior (Voa) nucleus (Voa and Vop have been collectivelytermed the ventral lateral anterior nucleus (VLa)); (3) ventrooralisposterior (Vop) nucleus; (4) subthalamic nuclei (campotomy), and; (5)CM-Pf thalamic nuclei. Generally, the ventrolateral thalamus has beenthe surgical target of choice in the treatment of Parkinson's diseaseand other systemically administered, drug resistant tremors. Brophy, B.P., et al., Thalamotomy for Parkinsonian Tremor, Stereotact FunctNeurosurg, 69;1-4:1997. Thalamic excitation of the cortex is necessaryfor almost all cortical activity.

Stereotactic surgery (assisted by neuroimaging and electrophysiologicrecordings) has been used in the management of advanced,pharmacoresistant Parkinson's disease, targeting hyperactive globuspallidus and subthalamic nuclei. An electrode or a probe is placed intothe brain using a brain atlas for reference with assistance from brainimaging by computer tomography or magnetic resonance imaging. Lesions indifferent parts of the pallidum (i.e. posteroventral pallidum), basalganglia, thalamus and subthalamic nuclei have been carried out to treatmotor disorders of Parkinson's disease. Unfortunately, surgical brainlesions create a risk of impairment to speech, visual and cognitivebrain areas. Neurotransplantation shows promise but requires furtherinvestigation. Additionally, deep brain stimulation using electrodes forthe suppression of tremor using can create problems due to wire erosion,lead friction, infection of the implantable pulse generator, malfunctionof the implantable pulse generator, electrical shock and lead migration.Other complications due to electrode stimulation can include dysarthria,disequilibrium, paresis and gait disorder. See e.g. Koller, W. C. etal., Surgical Treatment of Parkinson's Disease, J Neurol Sci167;1-10:1999, and Schuurman P. R., et al., A Comparison of ContinuousThalamic Stimulation and Thalamotomy for Suppression of Severe Tremor,NEJM 342(7);461-468:2000.

Aside from surgical ablation or stimulation, external radiotherapy(Gamma Knife Radiosurgery) has also been used to a limited extent forthe treatment of drug resistant parkinsonian tremors. Drawbacks withthis procedure are that the reduction in tremor is delayed by betweenone week and eight months after the radiosurgery, and that long termbenefits as well as radiation side effects are currently unknown.

As set forth, treatment of parkinsonian tremor and other movementdisorders has been carried out by thalamotomy and/or interruption ofpallidofugal fibers in the subthalamic region and pallidotomy has alsobeen used. Current concepts of basal ganglia circuitry propose that theloss of striatal dopamine in Parkinson's disease leads to overactivityof the striatal projection to the lateral segment of the globuspallidus. The resulting decrease in lateral pallidal activity results indisinhibition of the subthalamic nucleus, its main projection site.Increased subthalamic activity in turn causes overactivity of theinternal segment of the globus pallidus, which projects to thepedunculopontine nucleus (PPN) and the ventrolateral (VL) thalamus.Thus, overactivity in the subthalamic nucleus and internal pallidumproduces the parkinsonian symptoms of tremor, bradykinesia andhypokinesia through projections to the PPN and VL thalamus. Lesion inthe subthalamic nucleus and the results of pallidotomy, particularlyposteroventral pallidotomy, have permitted effective treatment ofakinesia in parkinsonian patients.

Botulinum Toxin

The genus Clostridium has more than one hundred and twenty sevenspecies, grouped according to their morphology and functions. Theanaerobic, gram positive bacterium Clostridium botulinum produces apotent polypeptide neurotoxin, botulinum toxin, which causes aneuroparalytic illness in humans and animals referred to as botulism.The spores of Clostridium botulinum are found in soil and can grow inimproperly sterilized and sealed food containers of home basedcanneries, which are the cause of many of the cases of botulism. Theeffects of botulism typically appear 18 to 36 hours after eating thefoodstuffs infected with a Clostridium botulinum culture or spores. Thebotulinum toxin can apparently pass unattenuated through the lining ofthe gut and attack peripheral motor neurons. Symptoms of botulinum toxinintoxication can progress from difficulty walking, swallowing, andspeaking to paralysis of the respiratory muscles and death.

Botulinum toxin type A is the most lethal natural biological agent knownto man. About 50 picograms of a commercially available botulinum toxintype A (purified neurotoxin complex)¹ is a LD₅₀ in mice (i.e. 1 unit).One unit of BOTOX® contains about 50 picograms (about 56 attomoles) ofbotulinum toxin type A complex. Interestingly, on a molar basis,botulinum toxin type A is about 1.8 billion times more lethal thandiphtheria, about 600 million times more lethal than sodium cyanide,about 30 million times more lethal than cobra toxin and about 12 milliontimes more lethal than cholera. Singh, Critical Aspects of BacterialProtein Toxins, pages 63-84 (chapter 4) of Natural Toxins II, edited byB. R. Singh et al., Plenum Press, New York (1996) (where the stated LD₅₀of botulinum toxin type A of 0.3 ng equals 1 U is corrected for the factthat about 0.05 ng of BOTOX® equals 1 unit). One unit (U) of botulinumtoxin is defined as the LD₅₀ upon intraperitoneal injection into femaleSwiss Webster mice weighing 18 to 20 grams each.

¹Available from Allergan, Inc., of Irvine, Calif. under the tradenameBOTOX® in 100 unit vials)

Seven immunologically distinct botulinum neurotoxins have beencharacterized, these being respectively botulinum neurotoxin serotypesA, B, C₁, D, E, F and G each of which is distinguished by neutralizationwith type-specific antibodies. The different serotypes of botulinumtoxin vary in the animal species that they affect and in the severityand duration of the paralysis they evoke. For example, it has beendetermined that botulinum toxin type A is 500 times more potent, asmeasured by the rate of paralysis produced in the rat, than is botulinumtoxin type B. Additionally, botulinum toxin type B has been determinedto be non-toxic in primates at a dose of 480 U/kg which is about 12times the primate LD₅₀ for botulinum toxin type A. Moyer E et al.,Botulinum Toxin Type B: Experimental and Clinical Experience, beingchapter 6, pages 71-85 of “Therapy With Botulinum Toxin”, edited byJankovic, J. et al. (1994), Marcel Dekker, Inc. Botulinum toxinapparently binds with high affinity to cholinergic motor neurons, istranslocated into the neuron and blocks the release of acetylcholine.

Regardless of serotype, the molecular mechanism of toxin intoxicationappears to be similar and to involve at least three steps or stages. Inthe first step of the process, the toxin binds to the presynapticmembrane of the target neuron through a specific interaction between theheavy chain, H chain, and a cell surface receptor; the receptor isthought to be different for each type of botulinum toxin and for tetanustoxin. The carboxyl end segment of the H chain, H_(C), appears to beimportant for targeting of the toxin to the cell surface.

In the second step, the toxin crosses the plasma membrane of thepoisoned cell. The toxin is first engulfed by the cell throughreceptor-mediated endocytosis, and an endosome containing the toxin isformed. The toxin then escapes the endosome into the cytoplasm of thecell. This step is thought to be mediated by the amino end segment ofthe H chain, H_(N), which triggers a conformational change of the toxinin response to a pH of about 5.5 or lower. Endosomes are known topossess a proton pump which decreases intra-endosomal pH. Theconformational shift exposes hydrophobic residues in the toxin, whichpermits the toxin to embed itself in the endosomal membrane. The toxin(or at a minimum the light chain) then translocates through theendosomal membrane into the cytoplasm.

The last step of the mechanism of botulinum toxin activity appears toinvolve reduction of the disulfide bond joining the heavy chain, Hchain, and the light chain, L chain. The entire toxic activity ofbotulinum and tetanus toxins is contained in the L chain of theholotoxin; the L chain is a zinc (Zn++) endopeptidase which selectivelycleaves proteins essential for recognition and docking ofneurotransmitter-containing vesicles with the cytoplasmic surface of theplasma membrane, and fusion of the vesicles with the plasma membrane.Tetanus neurotoxin, botulinum toxin types B, D, F, and G causedegradation of synaptobrevin (also called vesicle-associated membraneprotein (VAMP)), a synaptosomal membrane protein. Most of the VAMPpresent at the cytoplasmic surface of the synaptic vesicle is removed asa result of any one of these cleavage events. Botulinum toxin serotype Aand E cleave SNAP-25. Botulinum toxin serotype C₁ was originally thoughtto cleave syntaxin, but was found to cleave syntaxin and SNAP-25. Eachof the botulinum toxins specifically cleaves a different bond, exceptbotulinum toxin type B (and tetanus toxin) which cleave the same bond.

Botulinum toxins have been used in clinical settings for the treatmentof neuromuscular disorders characterized by hyperactive skeletalmuscles. A botulinum toxin type A complex has been approved by the U.S.Food and Drug Administration for the treatment of blepharospasm,strabismus and hemifacial spasm. Non-type A botulinum toxin serotypesapparently have a lower potency and/or a shorter duration of activity ascompared to botulinum toxin type A. Clinical effects of peripheralintramuscular botulinum toxin type A are usually seen within one week ofinjection. The typical duration of symptomatic relief from a singleintramuscular injection of botulinum toxin type A averages about threemonths.

Although all the botulinum toxins serotypes apparently inhibit releaseof the neurotransmitter acetylcholine at the neuromuscular junction,they do so by affecting different neurosecretory proteins and/orcleaving these proteins at different sites. For example, botulinum typesA and E both cleave the 25 kiloDalton (kD) synaptosomal associatedprotein (SNAP-25), but they target different amino acid sequences withinthis protein. Botulinum toxin types B, D, F and G act onvesicle-associated protein (VAMP, also called synaptobrevin), with eachserotype cleaving the protein at a different site. Finally, botulinumtoxin type C₁ has been shown to cleave both syntaxin and SNAP-25. Thesedifferences in mechanism of action may affect the relative potencyand/or duration of action of the various botulinum toxin serotypes.Apparently, a substrate for a botulinum toxin can be found in a varietyof different cell types. See e.g. Gonelle-Gispert, C., et al., SNAP-25aand -25b isoforms are both expressed in insulin-secreting cells and canfunction in insulin secretion, Biochem J. 1;339 (pt 1): 159-65:1999, andBoyd R. S. et al., The effect of botulinum neurotoxin-B on insulinrelease from a β-cell line, and Boyd R. S. et al., The insulin secretingβ-cell line, HIT-15, contains SNAP-25 which is a target for botulinumneurotoxin-A, both published at Mov Disord, 10(3):376:1995 (pancreaticislet B cells contains at least SNAP-25 and synaptobrevin).

The molecular weight of the botulinum toxin protein molecule, for allseven of the known botulinum toxin serotypes, is about 150 kD.Interestingly, the botulinum toxins are released by Clostridialbacterium as complexes comprising the 150 kD botulinum toxin proteinmolecule along with associated non-toxin proteins. Thus, the botulinumtoxin type A complex can be produced by Clostridial bacterium as 900 kD,500 kD and 300 kD forms. Botulinum toxin types B and C, is apparentlyproduced as only a 700 kD or 500 kD complex. Botulinum toxin type D isproduced as both 300 kD and 500 kD complexes. Finally, botulinum toxintypes E and F are produced as only approximately 300 kD complexes. Thecomplexes (i.e. molecular weight greater than about 150 kD) are believedto contain a non-toxin hemaglutinin protein and a non-toxin andnon-toxic nonhemaglutinin protein. These two non-toxin proteins (whichalong with the botulinum toxin molecule comprise the relevant neurotoxincomplex) may act to provide stability against denaturation to thebotulinum toxin molecule and protection against digestive acids whentoxin is ingested. Additionally, it is possible that the larger (greaterthan about 150 kD molecular weight) botulinum toxin complexes may resultin a slower rate of diffusion of the botulinum toxin away from a site ofintramuscular injection of a botulinum toxin complex.

In vitro studies have indicated that botulinum toxin inhibits potassiumcation induced release of both acetylcholine and norepinephrine fromprimary cell cultures of brainstem tissue. Additionally, it has beenreported that botulinum toxin inhibits the evoked release of bothglycine and glutamate in primary cultures of spinal cord neurons andthat in brain synaptosome preparations botulinum toxin inhibits therelease of each of the neurotransmitters acetylcholine, dopamine,norepinephrine (Habermann E., et al., Tetanus Toxin and Botulinum A andC Neurotoxins Inhibit Noradrenaline Release From Cultured Mouse Brain, JNeurochem 51(2);522-527:1988) CGRP, substance P and glutamate(Sanchez-Prieto, J., et al., Botulinum Toxin A Blocks GlutamateExocytosis From Guinea Pig Cerebral Cortical Synaptosomes, Eur J.Biochem 165;675-681:1987. Thus, when adequate concentrations are used,stimulus-evoked release of most neurotransmitters is blocked bybotulinum toxin. See e.g. Pearce, L. B., Pharmacologic Characterizationof Botulinum Toxin For Basic Science and Medicine, Toxicon35(9);1373-1412 at 1393; Bigalke H., et al., Botulinum A NeurotoxinInhibits Non-Cholinergic Synaptic Transmission in Mouse Spinal CordNeurons in Culture, Brain Research 360;318-324:1985; Habermann E.,Inhibition by Tetanus and Botulinum A Toxin of the release of [ ³H]Noradrenaline and [ ³ H]GABA From Rat Brain Homogenate, Experientia44;224-226:1988, Bigalke H., et al., Tetanus Toxin and Botulinum A ToxinInhibit Release and Uptake of Various Transmitters, as Studied withParticulate Preparations From Rat Brain and Spinal Cord,Naunyn-Schmiedeberg's Arch Pharmacol 316;244-251:1981, and; Jankovic J.et al., Therapy With Botulinum Toxin, Marcel Dekker, Inc., (1994), page5.

Botulinum toxin type A can be obtained by establishing and growingcultures of Clostridium botulinum in a fermenter and then harvesting andpurifying the fermented mixture in accordance with known procedures. Allthe botulinum toxin serotypes are initially synthesized as inactivesingle chain proteins which must be cleaved or nicked by proteases tobecome neuroactive. The bacterial strains that make botulinum toxinserotypes A and G possess endogenous proteases and serotypes A and G cantherefore be recovered from bacterial cultures in predominantly theiractive form. In contrast, botulinum toxin serotypes C₁, D and E aresynthesized by nonproteolytic strains and are therefore typicallyunactivated when recovered from culture. Serotypes B and F are producedby both proteolytic and nonproteolytic strains and therefore can berecovered in either the active or inactive form. However, even theproteolytic strains that produce, for example, the botulinum toxin typeB serotype only cleave a portion of the toxin produced. The exactproportion of nicked to unnicked molecules depends on the length ofincubation and the temperature of the culture. Therefore, a certainpercentage of any preparation of, for example, the botulinum toxin typeB toxin is likely to be inactive, possibly accounting for the knownsignificantly lower potency of botulinum toxin type B as compared tobotulinum toxin type A. The presence of inactive botulinum toxinmolecules in a clinical preparation will contribute to the overallprotein load of the preparation, which has been linked to increasedantigenicity, without contributing to its clinical efficacy.Additionally, it is known that botulinum toxin type B has, uponintramuscular injection, a shorter duration of activity and is also lesspotent than botulinum toxin type A at the same dose level.

High quality crystalline botulinum toxin type A can be produced from theHall A strain of Clostridium botulinum with characteristics of ≧3×10⁷U/mg, an A₂₆₀/A₂₇₈ of less than 0.60 and a distinct pattern of bandingon gel electrophoresis. The known Schantz process can be used to obtaincrystalline botulinum toxin type A, as set forth in Schantz, E. J., etal, Properties and use of Botulinum toxin and Other MicrobialNeurotoxins in Medicine, Microbiol Rev. 56;80-99:1992. Generally, thebotulinum toxin type A complex can be isolated and purified from ananaerobic fermentation by cultivating Clostridium botulinum type A in asuitable medium. The known process can also be used, upon separation outof the non-toxin proteins, to obtain pure botulinum toxins, such as forexample: purified botulinum toxin type A with an approximately 150 kDmolecular weight with a specific potency of 1-2×10⁸ LD₅₀ U/mg orgreater; purified botulinum toxin type B with an approximately 156 kDmolecular weight with a specific potency of 1-2×10⁸ LD₅₀ U/mg orgreater, and; purified botulinum toxin type F with an approximately 155kD molecular weight with a specific potency of 1-2×10⁷ LD₅₀ U/mg orgreater.

Botulinum toxins and/or botulinum toxin complexes can be obtained fromList Biological Laboratories, Inc., Campbell, Calif.; the Centre forApplied Microbiology and Research, Porton Down, U.K.; Wako (Osaka,Japan), Metabiologics (Madison, Wis.) as well as from Sigma Chemicals ofSt Louis, Mo.

Pure botulinum toxin is so labile that it is generally not used toprepare a pharmaceutical composition. Furthermore, the botulinum toxincomplexes, such a the toxin type A complex are also extremelysusceptible to denaturation due to surface denaturation, heat, andalkaline conditions. Inactivated toxin forms toxoid proteins which maybe immunogenic. The resulting antibodies can render a patient refractoryto toxin injection.

As with enzymes generally, the biological activities of the botulinumtoxins (which are intracellular peptidases) is dependant, at least inpart, upon their three dimensional conformation. Thus, botulinum toxintype A is detoxified by heat, various chemicals surface stretching andsurface drying. Additionally, it is known that dilution of the toxincomplex obtained by the known culturing, fermentation and purificationto the much, much lower toxin concentrations used for pharmaceuticalcomposition formulation results in rapid detoxification of the toxinunless a suitable stabilizing agent is present. Dilution of the toxinfrom milligram quantities to a solution containing nanograms permilliliter presents significant difficulties because of the rapid lossof specific toxicity upon such great dilution. Since the toxin may beused months or years after the toxin containing pharmaceuticalcomposition is formulated, the toxin can stabilized with a stabilizingagent such as albumin and gelatin.

A commercially available botulinum toxin containing pharmaceuticalcomposition is sold under the trademark BOTOX® (available from Allergan,Inc., of Irvine, Calif.). BOTOX® consists of a purified botulinum toxintype A complex, albumin and sodium chloride packaged in sterile,vacuum-dried form. The botulinum toxin type A is made from a culture ofthe Hall strain of Clostridium botulinum grown in a medium containingN—Z amine and yeast extract. The botulinum toxin type A complex ispurified from the culture solution by a series of acid precipitations toa crystalline complex consisting of the active high molecular weighttoxin protein and an associated hemagglutinin protein. The crystallinecomplex is re-dissolved in a solution containing saline and albumin andsterile filtered (0.2 microns) prior to vacuum-drying. The vacuum-driedproduct is stored in a freezer at or below −5° C. BOTOX® can bereconstituted with sterile, non-preserved saline prior to intramuscularinjection. Each vial of BOTOX® contains about 100 units (U) ofClostridium botulinum toxin type A purified neurotoxin complex, 0.5milligrams of human serum albumin and 0.9 milligrams of sodium chloridein a sterile, vacuum-dried form without a preservative.

To reconstitute vacuum-dried BOTOX®, sterile normal saline without apreservative; (0.9% Sodium Chloride Injection) is used by drawing up theproper amount of diluent in the appropriate size syringe. Since BOTOX®may be denatured by bubbling or similar violent agitation, the diluentis gently injected into the vial. For sterility reasons BOTOX® ispreferably administered within four hours after the vial is removed fromthe freezer and reconstituted. During these four hours, reconstitutedBOTOX® can be stored in a refrigerator at about 2° C. to about 8° C.Reconstituted, refrigerated BOTOX® has been reported to retain itspotency for at least about two weeks. Neurology, 48:249-53:1997.

It has been reported that botulinum toxin type A has been used inclinical settings as follows:

(1) about 75-125 units of BOTOX® per intramuscular injection (multiplemuscles) to treat cervical dystonia;

(2) 5-10 units of BOTOX® per intramuscular injection to treat glabellarlines (brow furrows) (5 units injected intramuscularly into the procerusmuscle and 10 units injected intramuscularly into each corrugatorsupercilii muscle);

(3) about 30-80 units of BOTOX® to treat constipation by intrasphincterinjection of the puborectalis muscle;

(4) about 1-5 units per muscle of intramuscularly injected BOTOX® totreat blepharospasm by injecting the lateral pre-tarsal orbicularisoculi muscle of the upper lid and the lateral pre-tarsal orbicularisoculi of the lower lid.

(5) to treat strabismus, extraocular muscles have been injectedintramuscularly with between about 1-5 units of BOTOX®, the amountinjected varying based upon both the size of the muscle to be injectedand the extent of muscle paralysis desired (i.e. amount of dioptercorrection desired).

(6) to treat upper limb spasticity following stroke by intramuscularinjections of BOTOX® into five different upper limb flexor muscles, asfollows:

(a) flexor digitorum profundus: 7.5 U to 30 U

(b) flexor digitorum sublimus: 7.5 U to 30 U

(c) flexor carpi ulnaris: 10 U to 40 U

(d) flexor carpi radialis: 15 U to 60 U

(e) biceps brachii: 50 U to 200 U. Each of the five indicated muscleshas been injected at the same treatment session, so that the patientreceives from 90 U to 360 U of upper limb flexor muscle BOTOX® byintramuscular injection at each treatment session.

(7) to treat migraine, pericranial injected (injected symmetrically intoglabellar, frontalis and temporalis muscles) injection of 25 U of BOTOX®has showed significant benefit as a prophylactic treatment of migrainecompared to vehicle as measured by decreased measures of migrainefrequency, maximal severity, associated vomiting and acute medicationuse over the three month period following the 25 U injection.

Additionally, intramuscular botulinum toxin has been used in thetreatment of tremor in patient's with Parkinson's disease, although ithas been reported that results have not been impressive. Marjama-Lyons,J., et al., Tremor-Predominant Parkinson's Disease, Drugs & Aging16(4);273-278:2000.

It is known that botulinum toxin type A can have an efficacy for up to12 months (Naumann M., et al., Botulinum toxin type A in the treatmentof focal, axillary and palmar hyperhidrosis and other hyperhidroticconditions, European J. Neurology 6 (Supp 4): S111-S115:1999), and insome circumstances for as long as 27 months. Ragona, R. M., et al.,Management of parotid sialocele with botulinum toxin, The Laryngoscope109:1344-1346:1999. However, the usual duration of an intramuscularinjection of Botox® is typically about 3 to 4 months.

The success of botulinum toxin type A to treat a variety of clinicalconditions has led to interest in other botulinum toxin serotypes. Astudy of two commercially available botulinum type A preparations(BOTOX® and DYSPORT®) and preparations of botulinum toxins type B and F(both obtained from Wako Chemicals, Japan) has been carried out todetermine local muscle weakening efficacy, safety and antigenicpotential. Botulinum toxin preparations were injected into the head ofthe right gastrocnemius muscle (0.5 to 200.0 units/kg) and muscleweakness was assessed using the mouse digit abduction scoring assay(DAS). ED₅₀ values were calculated from dose response curves. Additionalmice were given intramuscular injections to determine LD₅₀ doses. Thetherapeutic index was calculated as LD₅₀/ED₅₀. Separate groups of micereceived hind limb injections of BOTOX® (5.0 to 10.0 units/kg) orbotulinum toxin type B (50.0 to 400.0 units/kg), and were tested formuscle weakness and increased water consumption, the later being aputative model for dry mouth. Antigenic potential was assessed bymonthly intramuscular injections in rabbits (1.5 or 6.5 ng/kg forbotulinum toxin type B or 0.15 ng/kg for BOTOX®). Peak muscle weaknessand duration were dose related for all serotypes. DAS ED₅₀ values(units/kg) were as follows: BOTOX®: 6.7, DYSPORT®: 24.7, botulinum toxintype B: 27.0 to 244.0, botulinum toxin type F: 4.3. BOTOX® had a longerduration of action than botulinum toxin type B or botulinum toxin typeF. Therapeutic index values were as follows: BOTOX®: 10.5, DYSPORT®:6.3, botulinum toxin type B: 3.2. Water consumption was greater in miceinjected with botulinum toxin type B than with BOTOX®, althoughbotulinum toxin type B was less effective at weakening muscles. Afterfour months of injections 2 of 4 (where treated with 1.5 ng/kg) and 4 of4 (where treated with 6.5 ng/kg) rabbits developed antibodies againstbotulinum toxin type B. In a separate study, 0 of 9 BOTOX® treatedrabbits demonstrated antibodies against botulinum toxin type A. DASresults indicate relative peak potencies of botulinum toxin type A beingequal to botulinum toxin type F, and botulinum toxin type F beinggreater than botulinum toxin type B. With regard to duration of effect,botulinum toxin type A was greater than botulinum toxin type B, andbotulinum toxin type B duration of effect was greater than botulinumtoxin type F. As shown by the therapeutic index values, the twocommercial preparations of botulinum toxin type A (BOTOX® and DYSPORT®)are different. The increased water consumption behavior observedfollowing hind limb injection of botulinum toxin type B indicates thatclinically significant amounts of this serotype entered the murinesystemic circulation. The results also indicate that in order to achieveefficacy comparable to botulinum toxin type A, it is necessary toincrease doses of the other serotypes examined. Increased dosage cancomprise safety. Furthermore, in rabbits, type B was more antigenic thanwas BOTOX®, possibly because of the higher protein load injected toachieve an effective dose of botulinum toxin type B. Aoki, K. R.,Preclinical update on BOTOX(botulinum toxin type A)-purified neurotoxincomplex relative to other botulinum neurotoxin preparations, Eur JNeurol 1999 Nov;6(Suppl 4):S3-S10.

In addition to having pharmacologic actions at the peripheral location,botulinum toxins may also have inhibitory effects in the central nervoussystem. Work by Weigand et al, (¹²⁵ I-labelled botulinum Aneurotoxin:pharmacokinetics in cats after intramuscular injection,Nauny-Schmiedeberg's Arch. Pharmacol. 1976; 292, 161-165), andHabermann, (¹²⁵ I-labelled Neurotoxin from clostridium botulinumA:preperation, binding to synaptosomes and ascent to the spinal cord,Nauny-Schmiedeberg's Arch. Pharmacol. 1974; 281, 47-56) showed thatbotulinum toxin is able to ascend to the spinal area by retrogradetransport. As such, a botulinum toxin injected at a peripheral location,for example intramuscularly, may be retrograde transported to the spinalcord.

U.S. Pat. No. 5,989,545 discloses that a modified clostridial neurotoxinor fragment thereof, preferably a botulinum toxin, chemically conjugatedor recombinantly fused to a particular targeting moiety can be used totreat pain by administration of the agent to the spinal cord.

Acetylcholine

Typically only a single type of small molecule neurotransmitter isreleased by each type of neuron in the mammalian nervous system. Theneurotransmitter acetylcholine is secreted by neurons in many areas ofthe brain, but specifically by the large pyramidal cells of the motorcortex, by several different neurons in the basal ganglia, by the motorneurons that innervate the skeletal muscles, by the preganglionicneurons of the autonomic nervous system (both sympathetic andparasympathetic), by the postganglionic neurons of the parasympatheticnervous system, and by some of the postganglionic neurons of thesympathetic nervous system. Essentially, only the postganglionicsympathetic nerve fibers to the sweat glands, the piloerector musclesand a few blood vessels are cholinergic, as most of the postganglionicneurons of the sympathetic nervous system secret the neurotransmitternorepinephine. In most instances acetylcholine has an excitatory effect.However, acetylcholine is known to have inhibitory effects at some ofthe peripheral parasympathetic nerve endings, such as inhibition ofheart rate by the vagal nerve.

The efferent signals of the autonomic nervous system are transmitted tothe body through either the sympathetic nervous system or theparasympathetic nervous system. The preganglionic neurons of thesympathetic nervous system extend from preganglionic sympathetic neuroncell bodies located in the intermediolateral horn of the spinal cord.The preganglionic sympathetic nerve fibers, extending from the cellbody, synapse with postganglionic neurons located in either aparavertebral sympathetic ganglion or in a prevertebral ganglion. Since,the preganglionic neurons of both the sympathetic and parasympatheticnervous system are cholinergic, application of acetylcholine to theganglia will excite both sympathetic and parasympathetic postganglionicneurons.

Acetylcholine activates two types of receptors, muscarinic and nicotinicreceptors. The muscarinic receptors are found in all effector cellsstimulated by the postganglionic, neurons of the parasympathetic nervoussystem as well as in those stimulated by the postganglionic cholinergicneurons of the sympathetic nervous system. The nicotinic receptors arefound in the adrenal medulla, as well as within the autonomic ganglia,that is on the cell surface of the postganglionic neuron at the synapsebetween the preganglionic and postganglionic neurons of both thesympathetic and parasympathetic systems. Nicotinic receptors are alsofound in many nonautonomic nerve endings, for example in the membranesof skeletal muscle fibers at the neuromuscular junction.

Acetylcholine is released from cholinergic neurons when small, clear,intracellular vesicles fuse with the presynaptic neuronal cell membrane.A wide variety of non-neuronal secretory cells, such as, adrenal medulla(as well as the PC12 cell line) and pancreatic islet cells releasecatecholamines and parathyroid hormone, respectively, from largedense-core vesicles. The PC12 cell line is a clone of ratpheochromocytoma cells extensively used as a tissue culture model forstudies of sympathoadrenal development. Botulinum toxin inhibits therelease of both types of compounds from both types of cells in vitro,permeabilized (as by electroporation) or by direct injection of thetoxin into the denervated cell. Botulinum toxin is also known to blockrelease of the neurotransmitter glutamate from cortical synaptosomescell cultures.

A neuromuscular junction is formed in skeletal muscle by the proximityof axons to muscle cells. A signal transmitted through the nervoussystem results in an action potential at the terminal axon, withactivation of ion channels and resulting release of the neurotransmitteracetylcholine from intraneuronal synaptic vesicles, for example at themotor endplate of the neuromuscular junction. The acetylcholine crossesthe extracellular space to bind with acetylcholine receptor proteins onthe surface of the muscle end plate. Once sufficient binding hasoccurred, an action potential of the muscle cell causes specificmembrane ion channel changes, resulting in muscle cell contraction. Theacetylcholine is then released from the muscle cells and metabolized bycholinesterases in the extracellular space. The metabolites are recycledback into the terminal axon for reprocessing into further acetylcholine.

Cholinergic Brain Systems

Cholinergic influence of both the motor and visual thalamus originatesfrom both the brainstem and the basal forebrain. See e.g. Billet S., etal., Cholinergic Projections to the Visual Thalamus and SuperiorColliculus, Brain Res. 847;121-123:1999 and Oakman, S. A. et al.,Characterization of the Extent of Pontomesencephalic CholinergicNeurons' projections to the Thalamus: Comparison with Projections toMidbrain Dopaminergic Groups, Neurosci 94(2);529-547;1999. Thus, it isknown based on histochemical studies using acetylcholinesterase (AchE)staining and retrograde tracing with choline acetyltransferase (ChAT)immunochemistry that there can be ascending cholinergic stimulation bythe brainstem of thalamic neurons. Steriade M. et al., Brain CholinergicSystems, Oxford University Press (1990), chapter 1. Indeed, manythalamic nuclei receive dense cholinergic innervation from brainstemreticular formations. Ibid, page 167. Known brainstem cholinergic cellgroups are located within: (1) the rostral pons at what is termed a Ch5location, which is located within the central tegmental field around thebrachium conjunctivum, forming a pedunculopontine tegmental nucleus,and; (2) the caudal part of the midbrain, at what is termed a Ch6location, the laterodorsal tegmental nucleus, which is embedded in theperiaqueductal and periventricular gray matter. The Ch5 and Ch6 cellgroups can consist almost exclusively of cholinergic neurons andtogether form the pontine cholinergic system. The Ch5-Ch6 cholinergicgroups provide direct ascending projections that terminate in a numberof target structure in the midbrain, diencephalon and telencephalon,including the superior colliculus, anterior pretectal area, interstitialmagnocellular nucleus of the posterior commissure, lateral habenularnucleus, thalamus, magnocellular preoptic nucleus, lateral mammillarynucleus, basal forebrain, olfactory bulb, medial prefrontal cortex andpontine nuclei. Stone T. W., CNS Neurotransmitters and Neuromodulators:Acetylcholine, CRC Press (1995), page 16. See also Schafer M. K.-H. etal., Cholinergic Neurons and Terminal Fields Revealed by Immunochemistryfor the Vesicular Acetylcholine Transporter. I. Central Nervous System,Neuroscience, 84(2);331-359:1998. Three dimensional localization ofCh1-8 cholinergic nuclei have been mapped in humans. See e.g. Tracey, D.J., et al., Neurotransmitters in the Human Brain, Plenum Press (1995),pages 136-139.

Additionally, the basal forebrain (proencephalon) provides cholinergicinnervation of the dorsal thalamus, as well as to the neocortex,hippocampus, amygdala and olfactory bulb. See e.g. Steridae, page136-136, supra. Basal forebrain areas where the great proportion ofneurons are cholinergic include the medial septal nucleus (Ch1), thevertical branches of the diagonal band nuclei (Ch2), the horizontalbranches of the diagonal band nuclei (Ch3), and the magnocellularnucleus basalis (Ch4), which is located dorsolaterally to the Ch3 cellgroup. Ch1 and Ch2 provide the major component of cholinergic projectionto the hippocampus. The cells in the Ch3 sector project to the olfactorybulb.

Furthermore, cholinergic neurons are present in the thalamus. Rico, B.et al., A Population of Cholinergic Neurons is Present in the MacaqueMonkey Thalamus, Eur J Neurosci, 10;2346-2352:1998.

Abnormalities in the brain's cholinergic system have been consistentlyidentified in a variety of neuropsychiatric disorders includingAlzheimer's disease, Parkinson's disease and dementia with Lewy bodies.Thus, in Alzheimer's disease there is hypoactivity of cholinergicprojections to the hippocampus and cortex. In individuals with dementiawith Lewy bodies extensive neocortical cholinergic deficits are believedto exist and in Parkinson's disease there is a loss of pedunculopontinecholinergic neurons. Notably, in vivo imaging of cholinergic activity inthe human brain has been reported. Perry, et al., Acetylcholine in Mind:a Neurotransmitter Correlate of Consciousness?, TINS 22(6);273-280:1999.

What is needed therefore is a method for effectively treating a movementdisorder by administration of a pharmaceutical which has thecharacteristics of long duration of activity, low rates of diffusion outof a chosen intracranial target tissue where administered, and nominalsystemic effects at therapeutic dose levels.

SUMMARY

The present invention meets this need and provides methods foreffectively treating a movement disorder by intracranial administrationof a neurotoxin which has the characteristics of long duration ofactivity, low rates of diffusion out of an intracranial site whereadministered and insignificant systemic effects at therapeutic doselevels.

The following definitions apply herein:

“About” means approximately or nearly and in the context of a numericalvalue or range set forth herein means ±10% of the numerical value orrange recited or claimed.

“Biological activity” includes, with regard to a neurotoxin, the abilityto influence synthesis, exocytosis, receptor binding and/or uptake of aneurotransmitter, such as acetylcholine, or of an endocrine or exocrinesecretory product, such as insulin or pancreatic juice, respectively.

“Local administration” means direct administration of a pharmaceuticalat or to the vicinity of a site on or within an animal body, at whichsite a biological effect of the pharmaceutical is desired. Localadministration excludes systemic routes of administration, such asintravenous or oral administration.

“Neurotoxin” means a biologically active molecule with a specificaffinity for a neuronal cell surface receptor. Neurotoxin includesClostridial toxins both as pure toxin and as complexed with one to morenon-toxin, toxin associated proteins

“Intracranial” means within the cranium or at or near the dorsal end ofthe spinal cord and includes the medulla, brain stem, pons, cerebellumand cerebrum.

A method for treating a movement disorder within the scope of thepresent invention can be by intracranial administration of a neurotoxinto a patient to thereby alleviate a symptom of the movement disorder.The neurotoxin is made by a bacterium selected from the group consistingof Clostridium botulinum, Clostridium butyricum and Clostridium beratti,or can be expressed by a suitable host (i.e. a recombinantly altered E.coli) which encodes for a neurotoxin made by Clostridium botulinum,Clostridium butyricum or Clostridium beratti. Preferably, the neurotoxinis a botulinum toxin, such as a botulinum toxin type A, B, C₁, D, E, Fand G.

The neurotoxin can be administered to various brain areas fortherapeutic treatment of a movement disorder, including to a lower brainregion, to a pontine region, to a mesopontine region, to a globuspallidus and/or to a thalamic region of the brain.

The neurotoxin can be a modified neurotoxin, that is a neurotoxin whichhas at least one of its amino acids deleted, modified or replaced, ascompared to a native or the modified neurotoxin can be a recombinantproduced neurotoxin or a derivative or fragment thereof.

Intracranial administration of a neurotoxin according to the presentinvention can include the step of implantation of controlled releasebotulinum toxin system. A detailed embodiment of the present inventioncan be a method for treating a movement disorder by intracranialadministration of a therapeutically effective amount of a botulinumtoxin to a patient to thereby treating a symptom of a movement disorder.The movement disorders treated can include Parkinson's disease,Huntington's Chorea, progressive supranuclear palsy, Wilson's disease,Tourettes syndrome, epilepsy, chronic tremor, tics , dystonias andspasticity.

A further embodiment within the scope of the present invention can be amethod for treating a movement disorder, the method comprising the stepsof: selecting a neurotoxin with tremor suppressant activity; choosing anintracranial target tissue which influences a movement disorder; and;intracranially administering to the target tissue a therapeuticallyeffective amount of the neurotoxin selected, thereby treating themovement disorder.

Thus, a method for treating a movement disorder according to the presentinvention can have the step of intracranial administration of aneurotoxin to a mammal, thereby alleviating a symptom of a movementdisorder experienced by the mammal. Most preferably, the botulinum toxinused is botulinum toxin type A because of the high potency, readyavailability and long history of clinical use of botulinum toxin type Ato treat various disorders.

I have surprisingly found that a botulinum toxin, such as botulinumtoxin type A, can be intracranially administered in amounts betweenabout 10⁻³ U/kg and about 10 U/kg to alleviate a movement disorderexperienced by a human patient. Preferably, the botulinum toxin used isintracranially administered in an amount of between about 10⁻² U/kg andabout 1 U/kg. More preferably, the botulinum toxin is administered in anamount of between about 10⁻¹ U/kg and about 1 U/kg. Most preferably, thebotulinum toxin is administered in an amount of between about 0.1 unitand about 5 units. Significantly, the movement disorder alleviatingeffect of the present disclosed methods can persist for between about 2months to about 6 months when administration is of aqueous solution ofthe neurotoxin, and for up to about five years when the neurotoxin isadministered as a controlled release implant.

A further preferred method within the scope of the present invention isa method for treating a movement disorder by selecting a neurotoxin withtremor suppressant activity, choosing an intracranial target tissuewhich influences a movement disorder; and intracranially administeringto the target tissue a therapeutically effective amount of theneurotoxin selected.

Another preferred method within the scope of the present invention is amethod for improving patient function, the method comprising the step ofintracranially administering a neurotoxin to a patient, therebyimproving patient function as determined by improvement in one or moreof the factors of reduced pain, reduced time spent in bed, increasedambulation, healthier attitude and a more varied lifestyle.

DESCRIPTION

The present invention is based on the discovery that significant andlong lasting relief from a variety of different movement disorders canbe achieved by intracranial administration of a neurotoxin. Intracranialadministration permits the blood brain barrier to be bypassed anddelivers much more toxin to the brain than is possible by a systemicroute of administration. Furthermore, systemic administration of aneurotoxin, such as a botulinum toxin, is contraindicated due to thesevere complications (i.e. botulism) which can result from entry of abotulinum toxin into the general circulation. Additionally, sincebotulinum toxin does not penetrate the blood brain barrier to anysignificant extent, systemic administration of a botulinum toxin has nopractical application to treat an intracranial target tissue.

The present invention encompasses any suitable method for intracranialadministration of a neurotoxin to a selected target tissue, includinginjection of an aqueous solution of a neurotoxin and implantation of acontrolled release system, such as a neurotoxin incorporating polymericimplant at the selected target site. Use of a controlled release implantreduces the need for repeat injections.

Intracranial implants are known. For example, brachytherapy formalignant gliomas can include stereotactically implanted, temporary,iodine-125 interstitial catheters. Scharfen. C. O., et al., HighActivity Iodine - 125 Interstitial Implant For Gliomas, Int. J.Radiation Oncology Biol Phys 24(4);583-591:1992. Additionally,permanent, intracranial, low dose ¹²⁵I seeded catheter implants havebeen used to treat brain tumors. Gaspar, et al., Permanent ¹²⁵ IImplants for Recurrent Malignant Gliomas, Int J Radiation Oncology BiolPhys 43(5);977-982:1999. See also chapter 66, pages 577-580, BellezzaD., et al., Stereotactic Interstitial Brachytherapy, in Gildenberg P. L.et al., Textbook of Stereotactic and Functional Neurosurgery,McGraw-Hill (1998).

Furthermore, local administration of an anti cancer drug to treatmalignant gliomas by interstitial chemotherapy using surgicallyimplanted, biodegradable implants is known. For example, intracranialadministration of 3-bis(chloro-ethyl)-1-nitrosourea (BCNU) (Carmustine)containing polyanhydride wafers, has.found therapeutic application.Brem, H. et al., The Safety of Interstitial Chemotherapy withBCNU-Loaded Polymer Followed by Radiation Therapy in the Treatment ofNewly Diagnosed Malignant Gliomas: Phase I Trial, J Neuro-Oncology26:111-123:1995.

A polyanhydride polymer, GLIADEL® (Stolle R & D, Inc., Cincinnati, Ohio)a copolymer of poly-carboxyphenoxypropane and sebacic acid in a ratio of20:80 has been used to make implants, intracranially implanted to treatmalignant gliomas. Polymer and BCNU can be co-dissolved in methylenechloride and spray-dried into microspheres. The microspheres can then bepressed into discs 1.4 cm in diameter and 1.0 mm thick by compressionmolding, packaged in aluminum foil pouches under nitrogen atmosphere andsterilized by 2.2 megaRads of gamma irradiation. The polymer permitsrelease of carmustine over a 2-3 week period, although it can take morethan a year for the polymer to be largely degraded. Brem, H., et al,Placebo-Controlled Trial of Safety and Efficacy of IntraoperativeControlled Delivery by Biodegradable Polymers of Chemotherapy forRecurrent Gliomas, Lancet 345; 1008-1012:1995.

An implant can be prepared by mixing a desired amount of a stabilizedneurotoxin (such as non-reconstituted BOTOX®) into a solution of asuitable polymer dissolved in methylene chloride, at room temperature.The solution can then be transferred to a Petri dish and the methylenechloride evaporated in a vacuum desiccator. Depending upon the implantsize desired and hence the amount of incorporated neurotoxin, a suitableamount of the dried neurotoxin incorporating implant is compressed atabout 8000 p.s.i. for 5 seconds or at 3000 p.s.i. for 17 seconds in amold to form implant discs encapsulating the neurotoxin. See e.g. FungL. K. et al., Pharmacokinetics of Interstitial Delivery of Carmustine4-Hydroperoxycyclophosphamide and Paclitaxel From a BiodegradablePolymer Implant in the Monkey Brain, Cancer Research 58;672-684:1998.

Diffusion of biological activity of a botulinum toxin within a tissueappears to be a function of dose and can be graduated. Jankovic J., etal Therapy With Botulinum Toxin, Marcel Dekker, Inc., (1994), page 150.

Local, intracranial delivery of a neurotoxin, such as a botulinum toxin,can provide a high, local therapeutic level of the toxin and cansignificantly prevent the occurrence of any systemic toxicity since manyneurotoxins, such as the botulinum toxins are too large to cross theblood brain barrier. A controlled release polymer capable of long term,local delivery of a neurotoxin to an intracranial site can circumventthe restrictions imposed by systemic toxicity and the blood brainbarrier, and permit effective dosing of an intracranial target tissue. Asuitable implant, as set forth in co-pending U.S. patent applicationSer. No. 09/587250 entitled “Neurotoxin Implant”, allows the directintroduction of a chemotherapeutic agent to a brain target tissue via acontrolled release polymer. The implant polymers used are preferablyhydrophobic so as to protect the polymer incorporated neurotoxin fromwater induced decomposition until the toxin is released into the targettissue environment.

Local intracranial administration of a botulinum toxin, according to thepresent invention, by injection or implant to e.g. the cholinergicthalamus presents as a superior alternative to thalamotomy in themanagement of inter alia tremor associated with Parkinson's disease

A method within the scope of the present invention includes stereotacticplacement of a neurotoxin containing implant using theRiechert-Mundinger unit and the ZD (Zamorano-Dujovny) multipurposelocalizing unit. A contrast-enhanced computerized tomography (CT) scan,injecting 120 ml of omnipaque, 350 mg iodine/ml, with 2 mm slicethickness can allow three dimensional multiplanar treatment planning(STP, Fischer, Freiburg, Germany). This equipment permits planning onthe basis of magnetic resonance imaging studies, merging the CT and MRItarget information for clear target confirmation.

The Leksell stereotactic system (Downs Surgical, Inc., Decatur, Ga.)modified for use with a GE CT scanner (General Electric Company,Milwaukee, Wis.) as well as the Brown-Roberts-Wells (BRW) stereotacticsystem (Radionics, Burlington, Mass.) can be used for this purpose.Thus, on the morning of the implant, the annular base ring of the BRWstereotactic frame can be attached to the patient's skull. Serial CTsections can be obtained at 3 mm intervals though the (target tissue)region with a graphite rod localizer frame clamped to the base plate. Acomputerized treatment planning program can be run on a VAX 11/780computer (Digital Equipment Corporation, Maynard, Mass.) using CTcoordinates of the graphite rod images to map between CT space and BRWspace.

Within wishing to be bound by theory, a mechanism can be proposed forthe therapeutic effects of a method practiced according to the presentinvention. Thus, a neurotoxin, such as a botulinum toxin, can inhibitneuronal exocytosis of several different CNS neurotransmitters, inparticular acetylcholine. It is known that cholinergic neurons arepresent in the thalamus. Additionally, cholinergic nuclei exist in thebasal ganglia or in the basal forebrain, with protections to motor andsensory cerebral regions. Thus, target tissues for a method within thescope of the present invention can include neurotoxin induced,reversible denervation of intracranial motor areas (such as thethalamus) as well as brain cholinergic systems themselves (such as basalnuclei) which project to the intracranial motor areas. For example,injection or implantation of a neurotoxin to a cholinergicallyinnervated thalamic nuclei (such as Vim) can result in (1)downregulation of Vim activity due to the action of the toxin uponcholinergic terminals projecting into the thalamus from basal ganglia,and; (2) attenuation of thalamic output due to the action of the toxinupon thalamic somata, both cholinergic and noncholinergic, therebyproducing a chemical thalamotomy.

Preferably, a neurotoxin used to practice a method within the scope ofthe present invention is a botulinum toxin, such as one of the serotypeA, B, C, D, E, F or G botulinum toxins. Preferably, the botulinum toxinused is botulinum toxin type A, because of its high potency in humans,ready availability, and known use for the treatment of skeletal andsmooth muscle disorders when locally administered by intramuscularinjection. Botulinum toxin type B is a less preferred neurotoxin to usein the practice of the disclosed methods because type B is known to havea significantly lower potency and efficacy as compared, to type A, isnot readily available, and has a limited history of clinical use inhumans. Furthermore, the higher protein load with regard to type B cancause immunogenic reaction to occur with development of antibodies tothe type B neurotoxin.

The amount of a neurotoxin selected for intracranial administration to atarget tissue according to the present disclosed invention can be variedbased upon criteria such as the movement disorder being treated, itsseverity, the extent of brain tissue involvement or to be treated,solubility characteristics of the neurotoxin toxin chosen as well as theage, sex, weight and health of the patient. For example, the extent ofthe area of brain tissue influenced is believed to be proportional tothe volume of neurotoxin injected, while the quantity of the tremorsuppressant effect is, for most dose ranges, believed to be proportionalto the concentration of neurotoxin injected. Methods for determining theappropriate route of administration and dosage are generally determinedon a case by case basis by the attending physician. Such determinationsare routine to one of ordinary skill in the art (see for example,Harrison's Principles of Internal Medicine (1998), edited by AnthonyFauci et al., 14^(th) edition, published by McGraw Hill).

I have found that a neurotoxin, such as a botulinum toxin, can beintracranially administered according to the present disclosed methodsin amounts of between about 10⁻³ U/kg to about 10 U/kg. A dose of about10⁻³ U/kg can result in a tremor suppressant effect if delivered to asmall intracranial nuclei. Intracranial administration of less thanabout 10⁻³ U/kg does not result in a significant or lasting therapeuticresult. An intracranial dose of more than 10 U/kg of a neurotoxin, suchas a botulinum toxin, poses a significant risk of denervation of sensoryor desirable motor functions of neurons adjacent to the target.

A preferred range for intracranial administration of a botulinum toxin,such as botulinum toxin type A, so as to achieve a tremor suppressanteffect in the patient treated is from about 10⁻² U/kg to about 1 U/kg.Less than about 10⁻² U/kg can result in a relatively minor, though stillobservable, tremor suppressant effect. A more preferred range forintracranial administration of a botulinum toxin, such as botulinumtoxin type A, so as to achieve an antinociceptive effect in the patienttreated is from about 10⁻¹ U/kg to about 1 U/kg. Less than about 10⁻¹U/kg can result in the desired therapeutic effect being of less than theoptimal or longest possible duration. A most preferred range forintracranial administration of a botulinum toxin, such as botulinumtoxin type A, so as to achieve a desired tremor suppressant effect inthe patient treated is from about 0.1 units to about 100 units.Intracranial administration of a botulinum toxin, such as botulinumtoxin type A, in this preferred range can provide dramatic therapeuticsuccess.

The present invention includes within its scope the use of anyneurotoxin which has a long duration tremor suppressant effect whenlocally applied intracranially to the patient. For example, neurotoxinsmade by any of the species of the toxin producing Clostridium bacteria,such as Clostridium botulinum, Clostridium butyricum, and Clostridiumberatti can be used or adapted for use in the methods of the presentinvention. Additionally, all of the botulinum serotypes A, B, C₁, D, E,F and G can be advantageously used in the practice of the presentinvention, although type A is the most preferred and type B the leastpreferred serotype, as explained above. Practice of the presentinvention can provide a tremor suppressant effect, per injection, for 3months or longer in humans.

Significantly, a method within the scope of the present invention canprovide improved patient function. “Improved patient function” can bedefined as an improvement measured by factors such as a reduced pain,reduced time spent in bed, increased ambulation, healthier attitude,more varied lifestyle and/or healing permitted by normal muscle tone.Improved patient function is synonymous with an improved quality of life(QOL). QOL can be assesses using, for example, the known SF-12 or SF-36health survey scoring procedures. SF-36 assesses a patient's physicaland mental health in the eight domains of physical functioning, rolelimitations due to physical problems, social functioning, bodily pain,general mental health, role limitations due to emotional problems,vitality, and general health perceptions. Scores obtained can becompared to published values available for various general and patientpopulations.

As set forth above, I have discovered that a surprisingly effective andlong lasting treatment of a movement disorder can be achieved byintracranial administration of a neurotoxin to an afflicted patient. Inits most preferred embodiment, the present invention is practiced byintracranial injection or implantation of botulinum toxin type A.

The present invention does include within its scope: (a) neurotoxinobtained or processed by bacterial culturing, toxin extraction,concentration, preservation, freeze drying and/or reconstitution and;(b) modified or recombinant neurotoxin, that is neurotoxin that has hadone or more amino acids or amino acid sequences deliberately deleted,modified or replaced by known chemical/biochemical amino acidmodification procedures or by use of known host cell/recombinant vectorrecombinant technologies, as well as derivatives or fragments ofneurotoxins so made.

Botulinum toxins for use according to the present invention can bestored in lyophilized, vacuum dried form in containers under vacuumpressure or as stable liquids. Prior to lyophilization the botulinumtoxin can be combined with pharmaceutically acceptable excipients,stabilizers and/or carriers, such as albumin. The lyophilized materialcan be reconstituted with saline or water.

EXAMPLES

The following examples set forth specific methods encompassed by thepresent invention to treat a movement disorder and are not intended tolimit the scope of the invention.

Example 1 Intracranial Target Tissue Localization and Methodology

Stereotactic procedures can be used for precise intracranialadministration of neurotoxin in aqueous form or as an implant to desiredtarget tissue. Thus, intracranial administration of a neurotoxin totreat a drug resistant tremor (i.e. a resting tremor, such as can occurin Parkinson's disease, or an action tremor, such as essential tremor),multiple sclerosis tremors, post traumatic tremors, post hemiplegictremors (post stroke spasticity), tremors associated with neuropathy,writing tremors and epilepsy can be carried out as follows.

A preliminary MRI scan of the patient can be carried out to obtain thelength of the anterior commissure-posterior commissure line and itsorientation to external bony landmarks. The base of the frame can thenbe aligned to the plane of the anterior commissure-posterior commissureline. CT guidance is used and can be supplemented with ventriculography.The posterior commissure can be visualized on 2mm CT slices and used asa reference point. Where the target injection site is the basal part ofthe ventral intermedius nucleus of the ventrolateral thalamus, averagecoordinates are 6.5 mm anterior to the posterior commissure, 11 mmlateral to the third ventricular wall and 2 mm above the anteriorcommissure-posterior commissure line. This location is not expected toencroach on the sensory thalamus or on a subthalamic region.

Physiological corroboration of target tissue localization can be by useof high and low frequency stimulation through an electrode accompanyingor incorporated into the long needle syringe used. A thermistorelectrode 1.6 mm in diameter with a 2 mm exposed tip can be used(Radionics, Burlington, Massachusetts). With electrode high frequencystimulation (75 Hz) paraesthetic responses can be elicited in theforearm and hand at 0.5-1.0 V using a Radionics lesion generator(Radionics Radiofrequency Lesion Generator Model RFG3AV). At lowfrequency (5 Hz) activation or disruption of tremor in the affected limboccurred at 2-3 V. With the methods of the present invention, theelectrode is not used to create a lesion. Following confirmation oftarget tissue localization, a neurotoxin can be injected, therebycausing a reversible, chemical thalamotomy. A typical injection is thedesired number of units (i.e. about 0.1 to about 5 units of a botulinumtoxin type A complex in about 0.1 ml to about 0.5 ml of water or saline.A low injection volume can be uses to minimize toxin diffusion away fromtarget. Typically, the neurotransmitter release inhibition effect can beexpected to wear off within about 2-4 months. Thus, an alternateneurotoxin format, neurotoxin incorporated within a polymeric implant,can be used to provide controlled, continuous release of therapeuticamount of the toxin at the desired location over a prolongedperiod.(i.e. from about 1 year to about 6 years), thereby obviating theneed for repeated toxin injections.

Several methods can be used for stereotactically guided injection of aneurotoxin to various intracranial targets, such as the subthalamicnucleus (STN) for treatment of Parkinson's disease (Parkinson'sdisease). Thus a stereotactic magnetic resonance (MRI) method relying onthree-dimensional (3D) T1-weighted images for surgical planning andmultiplanar T2-weighted images for direct visualization of the STN,coupled with electrophysiological recording and injection guidance forunilateral or bilateral STN injection can be used. See e.g. Bejjani, B.P., et al., Bilateral Subthalamic Stimulation for Parkinson's Disease byUsing Three-Dimensional Stereotactic Magnetic Resonance Imaging andElectrophysiological Guidance, J Neurosurg 92(4);615-25:2000. The STNscan be visualized as 3D ovoid biconvex hypointense structures located inthe upper mesencephalon. The coordinates of the centers of the STNs canbe determined with reference to the patient's anteriorcommissure-posterior commissure line by using as a landmark, theanterior border of the red nucleus.

Electrophysiological monitoring through several parallel tracks can beperformed simultaneously to define the functional target accurately.Microelectrode recording can identify high-frequency, spontaneous,movement-related activity and tremor-related cells within the STNs.Neurotoxin injection into the STN can improve contralateral rigidity andakinesia and suppress tremor when present. The central track, which isdirected at the predetermined target by using MRI imaging, can beselected for neurotoxin injection. No surgical complications areexpected. The patient can show significantly improved parkinsonian motordisability in the “off” and “on” medication states and use ofantiparkinsonian drug treatment can be dramatically reduced as is theseverity of levodopa-induced dyskinesias and motor fluctuations.

Computer-aided atlas-based functional neurosurgery methodology can beused to accurately and precisely inject the desired neurotoxin orimplant a neurotoxin controlled release implant. Such methodologiespermit three-dimensional display and real-time manipulation of cerebralstructures. Neurosurgical planning with mutually preregistered multiplebrain atlases in all three orthogonal orientations is therefore possibleand permits increased accuracy of target definition for neurotoxininjection or implantation, reduced time of the surgical procedure bydecreasing the number of tracts, and facilitates planning of moresophisticated trajectories. See e.g. Nowinski W. L. et al.,Computer-Aided Stereotactic Functional Neurosurgery Enhanced by the Useof the Multiple Brain Atlas Database, IEEE Trans Med Imaging19(1);62-69:2000.

Example 2 Treatment of Parkinson's Disease With Botulinum Toxin Type A

A 64 year old right-handed male presents with pronounced tremor of theextremities, bradykinesia, rigidity and postural changes such that hefrequently falls. A prominent pill rolling tremor is noted in his righthand. Stroke is ruled out and it is noted that the symptoms are worse onhis right side. Diagnosis of Parkinson's disease is made. Using CAT scanor MRI assisted stereotaxis, as set forth in Example 1 above, 2 units ofa botulinum toxin type A (such as BOTOX® or about 8 units of DYSPORT®)is injected into the left side of the globus pallidus. The patient isdischarged within 48 hours and with a few (1-7) days enjoys significantimprovement of the parkinsonian motor symptoms more clearly on theright, but also on his left side. His dyskinesias almost completelydisappear. The motor disorder symptoms of Parkinson's disease remainsignificantly alleviated for between about 2 to about 6 months. Forextended therapeutic relief, one or more polymeric implantsincorporating a suitable quantity of a botulinum toxin type A can beplaced at the target tissue site.

Example 3 Treatment of Parkinson's Disease With Botulinum Toxin Type B

A 68 year left handed male presents with pronounced tremor of theextremities, bradykinesia, rigidity and postural changes such that hefrequently falls. A prominent pill rolling tremor is noted on his leftside. Stroke is ruled out and it is noted that the symptoms are worse onhis left side. Diagnosis of Parkinson's disease is made. Using CAT scanor MRI assisted stereotaxis, as set forth in Example 1 above, from 10 toabout 50 units of a botulinum toxin type B preparation (such asNEUROBLOC® or INERVATE™) is injected into the right side of the globuspallidus. The patient is discharged within 48 hours and with a few (1-7)days enjoys significant improvement of the parkinsonian motor symptomsmore clearly on the left, but also on his right side. His dyskinesiasalmost completely disappear. The motor disorder symptoms of Parkinson'sdisease remain significantly alleviated for between about 2 to about 6months. For extended therapeutic relief, one or more polymeric implantsincorporating a suitable quantity of a botulinum toxin type B can beplaced at the target tissue site.

Example 4 Treatment of Parkinson's Disease With Botulinum Toxin TypesC₁-G

A female aged 71 is admitted with uncontrollable and frequent tremor.From 0.1 to 100 units of a botulinum toxin type C₁, D, E, F or G isinjected unilaterally Into the ventrolateral thalamus for the disablingtremors. CAT scan or MRI assisted stereotaxis, as set forth in Example 1above, supplemented by ventriculography is used. The patient isdischarged within 48 hours and with a few (1-7) days enjoys significantremission of tremors which remain significantly alleviated for betweenabout 2 to about 6 months. For extended therapeutic relief, one or morepolymeric implants incorporating a suitable quantity of a botulinumtoxin type C₁, D, E, F or G can be placed at the target tissue site.

Example 5 Treatment of Dystonia With Botulinum Toxin Type A

A 16 year old male child with severe, incapacitating dystonia, secondaryto cranial trauma, affecting the proximal limb muscles is a candidatefor unilateral thalamotomy on the left side, bilateral thalamotomycarrying a high risk of iatrogenic dysarthria and pseudubulbar effects.The patient has failed to respond or has become unresponsive totranscutaneous nerve stimulation, feedback display of the EMG andanticholinergics. The dystonia is relatively stable, the patient issufficiently fit to withstand surgery and is significantly disabled withdistal phasic and tonic limb dystonia.

A suitable stereotactic frame can be applied to the head with localanesthetic and ventriculography and stereotactic MRI can be performed.The stereotactic coordinates of the anterior commissure (AC) and theposterior commissure (PC) can be determined by using the computersoftware in the scanner. PC based software can be used to redraw thesagittal brain maps from the Schaltenbrand and Bailey and Schaltenbrandand Wahren atlases, stretched or shrunk as needed to the AC-PC distanceof the patient and ruled in stereotactic coordinates for the actualapplication of the frame to the patient's head. The target sites areselected, their coordinates are read off and appropriate frame settingsare made. A burr hole or twist -drill hole can be made at or rostral tothe coronal suture in the same sagittal plane as the target. This canfacilitate plotting the physiological data used for target corroborationsince the electrode trajectories traverse a single sagittal plane. Theventrocaudal nucleus of the thalamus (Vc) can be selected as aphysiological landmark, lying 15 mm from the midline. The Vc can beeasily recognized by recording individual tactile cells within it withtheir discrete receptive fields or by inducing paresthesias withstimulation in discreet projected fields.

A microelectrode recording needle (such a used for single fiberelectromyographic recording having an approximately 25 micron diameterrecording electrode) can be located within the bore of a microsyringeand is advanced toward the expected tactile representation of thefingers in the Vc and continuous recording is carried out to search foridentifiable neurons. Microstimulation can be performed everymillimeter, beginning about 10 mm above and extending to a variabledistance below the target. If the first microelectrode trajectoryenters, for example, the tactile representation of the lips of a patientwith upper limb dystonia, a second trajectory can be carried out 2 mmmore lateral. Upon encountering lower limb responses, the nexttrajectory can be made 2 mm more medial. Once the tactile representationof the hand is found, the next trajectory can be made 2 mm rostral toit, where recording reveals kinesthetic neurons that respond to bendingof specific contralateral joints or pressure on specific contralateralsites. If dystonia is confined to the leg, the process described abovecan be aimed at the thalamic representation for the leg.

Upon microstimulation localization of the stereotactically -MRI guidedrecording/stimulating needle electrode to the target, a neurotoxinimplant can be injected. The implant can comprise a neurotoxin, such asa of botulinum toxin type A, incorporated within biodegradable polymericmicrospheres or a biodegradable pellet, either implant format containingabout 20 total units (about 1 ng) of the toxin with implantcharacteristics of continuous release over a period of at least aboutfour years of a therapeutic level of the toxin at point of the implantrelease site and for a radius of about 2-3 mm on each side o the targetsite. The implant can release about 1 unit of toxin essentiallyimmediately and further amounts of about one unit cumulatively oversubsequent 2-4 months periods.

The patient's dystonic contractions can subside almost immediately, andcan remain substantially alleviated for between about 2 months to about6 months per toxin injection or for between about 1 to 5 years dependingupon the particular release characteristics of the implant polymer andthe quantity of neurotoxin loaded therein.

Example 6 Treatment of Dystonia With Botulinum Toxin Types B-G

The patient of example 5 above can be equivalently treated using thesame protocol and approach to target with between about 1 unit and about1000 units of a botulinum toxin type B, C₁, D, E, F or G in aqueoussolution or in the form of a suitable neurotoxin implant. With such atreatment, the dystonic contractions subside within 1-7 days, and remainsubstantially alleviated for between about 2-6 months per toxininjection or for between about 1 to 5 years depending upon theparticular release characteristics of the implant polymer and thequantity of neurotoxin loaded therein.

Example 7 Treatment of Tremor With Botulinum Toxin Type A

A 44 year old male presents with severe incapacitating tremor of threeyears duration which disrupts his activities of daily living. There isalso asymmetry of the motor symptoms between the two side of the bodyand levodopa has induced dyskinesia in the extremities. Tremor cells areidentified by stereotactic examination of the effect upon the tremor byelectrical stimulation of the proposed target cell. The effect ofstimulation is noted to inhibit the tremor. Stereotactic guided (as inExample 1) implant placement can be made at a site about 14 to 15 mmfrom the midline and 2-3 mm above the AC-PC line in the middle ofkinesthetic and/or voluntary tremor cells. The target site can be the VLor Vi.

The implant can be either an aqueous solution of botulinum toxin type Aincorporated within biodegradable polymeric microspheres or botulinumtoxin type A biodegradable pellet, either implant format containingabout 20 total units (about 1 ng) of the toxin with implantcharacteristics of continuous release over a period of at least aboutfour years of a therapeutic level of the toxin at point of the implantrelease site and in about 2-3 mm on each side. The implant can releaseabout 1 unit of toxin essentially immediately and further amounts ofabout one unit cumulatively over subsequent 2-4 months periods.

The patient's tremors can subside within 1-7 days, and can remainsubstantially alleviated for between about 2 months to about 6 monthsper toxin injection or for between about 1 to 5 years depending upon theparticular release characteristics of the implant polymer and thequantity of neurotoxin loaded therein. Notably, there can be significantattenuation of distal limb movements, both phasic and tonic on the rightside.

Example 8 Treatment of Tremor With Botulinum Toxin Types B-G

The patient of example 7 above can be equivalently treated using thesame protocol and approach to target with between about 1 unit and about1000 units of a botulinum toxin type B, C₁, D, E, F or G in aqueoussolution or in the form of a suitable neurotoxin implant. With such atreatment, the tremors can subside within 1-7 days, and can remainsubstantially alleviated for between about 2-6 months per toxininjection or for between about 1 to 5 years depending upon theparticular release characteristics of the implant polymer and thequantity of neurotoxin loaded therein.

Example 9 Treatment of Epilepsy With Botulinum Toxin Type A

A right handed, female patient age 22 presents with a history ofepilepsy. Based upon MRI and a study of EEG recording, a diagnosis oftemporal lobe epilepsy is made. An implant which provides about 5-50units of a neurotoxin (such as a botulinum toxin type A) can be insertedat the anterior part of the temporal lobe, 5-6 cm from the tip of thelobe along the middle temporal gyrus with a unilateral approach to thenondominant, left hemisphere. The epileptic seizures can besubstantially reduced within about 1-7 days, and can remainsubstantially alleviated for between about 2 months to about 6 monthsper toxin injection or for between about 1 to 5 years depending upon theparticular release characteristics of the implant polymer and thequantity of neurotoxin loaded therein.

Example 10 Treatment of Epilepsy With Botulinum Toxin Types B-G

The patient of example 9 above can be equivalently treated using thesame protocol and approach to target with between about 1 unit and about1000 units of a botulinum toxin type B, C₁, D, E, F or G in aqueoussolution or in the form of a suitable neurotoxin implant. With such atreatment, the epileptic seizures can subside within 1-7 days, and canremain substantially alleviated for between about 2-6 months per toxininjection or for between about 1 to 5 years depending upon theparticular release characteristics of the implant polymer and thequantity of neurotoxin loaded therein.

It is concluded that neurotoxin injection or implantation of acontrolled release neurotoxin implant according to the methods of thepresent invention, with the aid of 3D MR imaging andelectrophysiological guidance, can be a safe and effective therapy forpatients suffering from various movement disorders, such as severe,advanced levodopa-responsive Parkinson's disease. Suitable patientsinclude those who have become largely if not entirely refractory tochemotherapy, typically oral L-dopa, prior to intracranial neurotoxinadministration as set forth herein.

A method according to the present invention can also be used diversemovement disorders, including essential tremor, multiple sclerosisrelated tremors, post traumatic tremors, post hemiplegic tremors,parkinsonian tremors and epilepsy.

An intracranial neurotoxin administration method for treating a movementdisorder according to the invention disclosed herein for has manybenefits and advantages, including the following:

1. the symptoms of a movement disorder can be dramatically reduced.

2. the symptoms of a movement disorder can be reduced for from about twoto about four months per injection of neurotoxin and for from about oneyear to about five years upon use of a controlled release neurotoxinimplant.

3. the injected or implanted neurotoxin exerts an intracranial targettissue site specific tremor suppressant effect.

4. the injected or implanted neurotoxin shows little or no tendency todiffuse or to be transported away from the intracranial injection orimplantation site.

5. few or no significant undesirable side effects occur fromintracranial injection or implantation of the neurotoxin.

6. the amount of neurotoxin injected intracranially can be considerablyless than the amount of the same neurotoxin required by other routes ofadministration (i.e. intramuscular, intrasphincter, oral or parenteral)to achieve a comparable tremor suppressant effect.

7. the tremor suppressant effects of the present methods can result inthe desirable side effects of greater patient mobility, a more positiveattitude, and an improved quality of life.

8. high, therapeutic doses of a neurotoxin can be delivered to anintracranial target tissue over a prolonged period without systemictoxicity.

Although the present invention has been described in detail with regardto certain preferred methods, other embodiments, versions, andmodifications within the scope of the present invention are possible.For example, a wide variety of neurotoxins can be effectively used inthe methods of the present invention. Additionally, the presentinvention includes intracranial administration methods wherein two ormore neurotoxins, such as two or more botulinum toxins, are administeredconcurrently or consecutively. For example, botulinum toxin type A canbe administered intracranially until a loss of clinical response orneutralizing antibodies develop, followed by administration of botulinumtoxin type B. Alternately, a combination of any two or more of thebotulinum serotypes A-G can be intracranially administered to controlthe onset and duration of the desired therapeutic result. Furthermore,non-neurotoxin compounds can be intracranially administered prior to,concurrently with or subsequent to administration of the neurotoxin toproved adjunct effect such as enhanced or a more rapid onset of tremorsuppression before the neurotoxin, such as a botulinum toxin, begins toexert its more long lasting tremor suppressant effect.

My invention also includes within its scope the use of a neurotoxin,such as a botulinum toxin, in the preparation of a medicament for thetreatment of a movement disorder, by intracranial administration of theneurotoxin.

All references, articles, patents, applications and publications setforth above are incorporated herein by reference in their entireties.

Accordingly, the spirit and scope of the following claims should not belimited to the descriptions of the preferred embodiments set forthabove.

I claim:
 1. A method for temporarily alleviating a motor disordersymptom of Parkinson's disease, the method comprising the step ofintracranial administration of a botulinum toxin to a patient, therebytemporarily alleviating a motor disorder symptom of Parkinson's disease.2. The method of claim 1, wherein the botulinum toxin is selected fromthe group consisting of botulinum toxin types A, B, C, D, E, F and G. 3.The method of claim 1, wherein the botulinum toxin is botulinum toxintype A.
 4. The method of claim 1, wherein the botulinum toxin isadministered in an amount of between 10⁻³ U/kg and 10 U/kg.
 5. Themethod of claim 1, wherein the alleviating effect persists for betweenabout 1 month and about 5 years.
 6. The method of claim 1, wherein thebotulinum toxin is administered to a lower brain region.
 7. The methodof claim 1, wherein the botulinum toxin is administered to a pontineregion.
 8. The method of claim 1, wherein the botulinum toxin isadministered a mesopontine region.
 9. The method of claim 1, wherein thebotulinum toxin is administered to a globus pallidus.
 10. The method ofclaim 1 wherein the botulinum toxin is administered to a thalamus. 11.The method of claim 1, wherein the botulinum toxin is a recombinantproduced botulinum toxin or a derivative or fragment thereof.
 12. Themethod of claim 1, wherein the intracranial administration stepcomprises implantation of a controlled release botulinum toxin system.13. A method for temporarily alleviating a motor disorder symptom ofParkinson's disease, the method comprising the step of intracranialadministration of a therapeutically effective amount of a botulinumtoxin A to a patient, thereby temporarily alleviating within one toseven days a motor disorder symptom of Parkinson's disease.